Anti-SIRP-alpha antibodies and methods of use thereof

Abstract

The invention provides anti-SIRPA antibodies, methods of generating such antibodies, and therapeutic uses and methods employing the antibodies.

What is claimed is: 
     
       1. An isolated anti-signal regulatory protein a (SIRPA) antibody, wherein the anti-SIRPA antibody comprises a heavy chain variable region (VH) that comprises a complementarity determining region 1 (CDR1) comprising the amino acid sequence of SEQ ID NO:9, a CDR2 comprising the amino acid sequence of SEQ ID NO:10, and a CDR3 comprising the amino acid sequence of SEQ ID NO:11; and a light chain variable region (VL) that comprises a CDR1 comprising the amino acid sequence of SEQ ID NO:6, a CDR2 comprising the amino acid sequence of SEQ ID NO:7, and a CDR3 comprising the amino acid sequence of SEQ ID NO:8. 
     
     
       2. The anti-SIRPA antibody of  claim 1 , wherein the antibody does not substantially block binding of CD47 to SIRPA expressed on cells, wherein binding of the antibody to SIRPA decreases the level of SIRPA expressed on the cell surface, and/or wherein the antibody binds to the D2 domain of SIRPA, or the D3 domain of SIRPA. 
     
     
       3. The anti-SIRPA antibody of  claim 1 , wherein the antibody competes with an antibody comprising a VH sequence comprising the amino acid sequence of SEQ ID NO:2 and a VL sequence comprising the amino acid sequence of SEQ ID NO:3. 
     
     
       4. The anti-SIRPA antibody of  claim 1 , wherein the VH comprises an amino acid sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to the amino acid sequence of hSB-3F9-H1 or hSB-3F9-H2. 
     
     
       5. The anti-SIRPA antibody of  claim 1 , wherein the VL comprises an amino acid sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to the amino acid sequence of hSB-3F9-L1, hSB-3F9-L2, or hsB-3F9-L3. 
     
     
       6. The isolated anti-SIRPA antibody of  claim 1 , comprising a VH region comprising the amino acid sequence of SEQ ID NO:2 and a VL region comprising the amino acid sequence of SEQ ID NO:3. 
     
     
       7. The anti-SIRPA antibody of  claim 1 , wherein the antibody is a monoclonal antibody. 
     
     
       8. The anti-SIRPA antibody of  claim 1 , wherein the antibody is a humanized antibody. 
     
     
       9. The anti-SIRPA antibody of  claim 1 , wherein the antibody is an Fab, Fab′, Fab′-SH, F(ab′)2, Fv or scFv fragment. 
     
     
       10. The anti-SIRPA antibody of  claim 1 , wherein the anti-SIRPA antibody has an IgG1, IgG2, IgG3, or IgG4 isotype. 
     
     
       11. The anti-SIRPA antibody of  claim 10 , wherein:
 (a) the anti-SIRPA antibody has a human IgG1 isotype and comprises one or more amino acid substitutions in the Fc region at a residue position selected from the group consisting of: N297A, D265A, D270A, L234A, L235A, G237A, P238D, L328E, E233D, G237D, H268D, P271G, A330R, C226S, C229S, E233P, L234V, L234F, L235E, P331S, S267E, L328F, A330L, M252Y, S254T, T256E, N297Q, P238S, P238A, A327Q, A327G, P329A, K322A, T394D, and any combination thereof, wherein the numbering of the residues is according to EU numbering, or comprises an amino acid deletion in the Fc region at a position corresponding to glycine 236; 
 (b) the anti-SIRPA antibody has an IgG1 isotype and comprises an IgG2 isotype heavy chain constant domain 1(CH1) and hinge region, optionally wherein the IgG2 isotype CH1 and hinge region comprises the amino acid sequence of ASTKGPSVFP LAPCSRSTSE STAALGCLVK DYFPEPVTVS WNSGALTSGVHTFPAVLQSS GLYSLSSVVT VPSSNFGTQT YTCNVDHKPS NTKVDKTVERKCCVECPPCP (SEQ ID NO:34), and optionally wherein the antibody Fc region comprises a S267E amino acid substitution, a L328F amino acid substitution, or both, and/or a N297A or N297Q amino acid substitution, wherein the numbering of the residues is according to EU numbering; 
 (c) the anti-SIRPA antibody has an IgG2 isotype and comprises one or more amino acid substitutions in the Fc region at a residue position selected from the group consisting of: P238S, V234A, G237A, H268A, H268Q, V309L, A330S, P331S, C214S, C232S, C233S, S267E, L328F, M252Y, S254T, T256E, H268E, N297A, N297Q, A330L, and any combination thereof, wherein the numbering of the residues is according to EU numbering; 
 (d) the anti-SIRPA antibody has a human IgG4 isotype and comprises one or more amino acid substitutions in the Fc region at a residue position selected from the group consisting of: L235A, G237A, S228P, L236E, S267E, E318A, L328F, M252Y, S254T, T256E, E233P, F234V, L234A/F234A, S228P, S241P, L248E, T394D, N297A, N297Q, L235E, and any combination thereof, wherein the numbering of the residues is according to EU numbering; or 
 (e) the anti-SIRPA antibody has a hybrid IgG2/4 isotype, and optionally wherein the antibody comprises an amino acid sequence comprising amino acids 118 to 260 of human IgG2 and amino acids 261 to 447 of human IgG4, wherein the numbering of the residues is according to EU numbering. 
 
     
     
       12. The anti-SIRPA antibody of  claim 1 , wherein the anti-SIRPA antibody is conjugated to a detectable marker, a toxin, or a therapeutic agent. 
     
     
       13. The anti-SIRPA antibody of  claim 12 , wherein the anti-SIRPA antibody is conjugated to a toxin selected from the group consisting of ricin, ricin A chain, doxorubicin, daunorubicin, a maytansinoid, taxol, ethidium bromide, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, dihydroxy anthracin dione, actinomycin, diphtheria toxin,  Pseudomonas  exotoxin (PE) A, PE40, abrin, abrin A chain, modeccin A chain, alpha sarcin, gelonin, mitogellin, retstrictocin, phenomycin, enomycin, curicin, crotin, calicheamicin,  Saponaria officinalis  inhibitor, glucocorticoid, auristatin, auromycin, yttrium, bismuth, combrestatin, duocarmycins, dolastatin, cc1065, and a cisplatin. 
     
     
       14. A pharmaceutical composition comprising an anti-SIRPA antibody of  claim 1  and a physiologically acceptable carrier. 
     
     
       15. The anti-SIRPA antibody of  claim 1 , wherein the VH comprises the amino acid sequence of hSB-3F9-H1 or hSB-3F9-H2. 
     
     
       16. The anti-SIRPA antibody of  claim 1 , wherein the VL comprises the amino acid sequence of hSB-3F9-L1, hSB-3F9-L2, or hsB-3F9-L3. 
     
     
       17. The anti-SIRPA antibody of  claim 1 , wherein the VH comprises the amino acid sequence of hSB-3F9-H1 or hSB-3F9-H2 and wherein the VL comprises the amino acid sequence of hSB-3F9-L1, hSB-3F9-L2, or hsB-3F9-L3. 
     
     
       18. The anti-SIRPA antibody of  claim 1 , wherein the VH comprises the amino acid sequence of SEQ ID NO: 2. 
     
     
       19. The anti-SIRPA antibody of  claim 1 , wherein the VL comprises the amino acid sequence of SEQ ID NO: 3. 
     
     
       20. The anti-SIRPA antibody of  claim 10 , wherein the anti-SIRPA antibody has a human IgG1 isotype and wherein the Fc region comprises substitution P331S; or substitutions L234A, L235A, and P331S; or substitutions S267E and L328F.